FAP-α is a Mr 95 kDa cell surface-bound type II transmembrane glycoprotein and is a member of the serine prolyl oligopeptidase family. Comparison of amino acid sequences indicate that FAP-α is essentially identical to seprase (Goldstein L A, et al., Biochim Biophys Acta 1997, 1361(1):11-19) and is closely related to DPP IV (dipeptidylpeptidase IV), also known as CD26, another type II integral membrane protein. (Morimoto C, et al., Immunol Rev 1998, 161:55-70). These exoproteases cleave NH2-terminal dipeptides from polypeptides with L-proline or L-alanine immediately following the N-terminal amino acid. FAP-α has been found to have collagenase activity in vitro. (Goldstein L A, et al., supra; Jones B, et al., Blood 2003, 102(5):1641-1648). In addition to various families of proteolytic enzymes, such as matrix or disintegrin metalloproteases that serve as major collagenases, peptidase activity of FAP-α contributes to extracellular matrix (“ECM”) degradation. (Park J E, et al., J Biol Chem 1999, 274(51):36505-36512; Ghersi G, et al., J Biol Chem 2002, 277(32):29211-29241). This is not only a fundamental property of normal tissue repair and remodeling, but is also involved in the pathological processes of invasive growth. This property correlates with the expression of FAP-α in granulation tissue of healing wounds (Grinnell F, J Cell Biol 1994, 124(4):401-404), desmoplastic reactions (Yen T W, et al., Surgery 2002, 131(2):129-134), and in more than 90% of human epithelial carcinomas. (Garin-Chesa P, et al., Proc Nat'l Acad Sci USA 1990, 87(18):7235-7239). FAP-α is mainly expressed on the surface of mesenchymal cells that are involved in epithelium-mesenchyme interactions contributing to tissue remodeling. Consistent with its mesenchymal origin, FAP-α is also occasionally expressed by bone and soft tissue sarcomas. (Rettig W J, et al., Proc Natl Acad Sci USA 1988, 85(9):31103114). Immunohistochemical staining of colorectal carcinomas and breast cancer (Park J E, et al., supra; Scanlan M J, et al., Proc Natl Acad Sci USA 1994, 91(12):5657-5661) confirmed the specific expression of FAP-α by tumor stroma fibroblasts but not by malignant cells themselves. (See also, Sappino A P, et al., Int J Cancer 1988, 41(5):707-712). Observations made during a clinical Phase I study, which examined the biodistribution of a humanized anti-FAP-α antibody in patients with advanced or metastatic PAP-a-positive cancer, a minor low-grade uptake in the knees and shoulders of three patients without clinical symptoms of arthritis, could have led to the conclusion that FAP-α might also be present in human joints. (Scott A M, et al., Clin Cancer Res 2003, 9(5):1639-1647). Unfortunately, no further explanation or discussion of this observation was presented. Further, it is unknown whether these patients were suffering from osteoarthritis. In contrast, resting fibrocytes in normal adult tissue generally lack detectable FAP-α expression. (Rettig W J, et al., supra; Garin-Chesa P, et al., Proc Natl Acad Sci USA 1990, 87(18):7235-7239).
Further information on FAP-α can be found in, e.g., U.S. Pat. Nos. 5,587,299; 5,767,242; 5,965,373; and 6,846,910, incorporated by reference in their entireties.
Rheumatoid arthritis (“RA”) is a chronic inflammatory disease of unknown etiology and characterized by hyperplasia and chronic inflammation of the synovial membranes that invade deeply into the articular cartilage and bone. Activated fibroblast-like synoviocytes (“FLS”) in the lining layer of the synovium are one of the dominant cells involved in pannus formation and are key players in joint destruction. (Firestein G S, Arthritis Rheum 1996, 39(11):1781-1790; Pap T, et al., Arthritis Rheum 2000, 43(11):2531-2536). Rheumatoid FLS have been shown to proliferate in an anchorage independent manner and express increased proliferation markers and matrix degrading enzymes when compared with FLS from patients with osteoarthritis (“OA”). (Qu Z, et al., Arthritis Rheum 1994, 37(2):212-220; Bucala R, et al., J Exp Med 1991, 173(3):569-574; Lafyatis R, et al., J Clin Invest 1989, 83(4):12671276). Expression of the CD44v7/8 epitope is linked to the proliferative behavior of FLS obtained from patients with RA, whereas expression of variants containing CD44v3 is linked with their increased invasive capacity (Croft D R, et al., Eur J Immunol 1997, 27(7):1680-1684; Wibulswas A, et al., Am J Pathol 2000, 157(6):2037-2044; Wibulswas A, et al., Arthritis Rheum 2002, 46(8):2059-2064). Matrix metalloproteases (“MMP”) have been shown to be essential for degradation of the articular matrix, with MMP-1 and MMP-13 being considered as important candidates for joint destruction in RA (Tomita T, et al., Arthritis Rheum 2002, 46(2):373-378; Westhoff C S, et al., Arthritis Rheum 1999, 42(7):1517-1527). Invasion of migratory fibroblasts into connective tissue, however requires cell surface serine proteases, as well as metallocollagenases. (Ghersi G, et al., supra). Among these exoproteases that may cooperate with interstitial collagenase are groups of serine prolyl-peptidases such as DPP IV/CD26 and FAP-α/seprase. (Park J E, et al., supra; De Meester I, et al., Immunol Today 1999, 20(8):367-375). Fibroblasts with smooth muscle differentiation, termed myofibroblasts, are generally accepted to be the main source for extracellular matrix degrading factors. Furthermore, FLS with a myofibroblast-like molecular phenotype have been identified in the intimal lining layer of inflamed rheumatoid synovium. (Kasperkovitz P V, et al., Arthritis Rheum 2005, 52(2):430-441).
Despite the elucidation of FLS as a key player in joint inflammation and proteolytic enzymes like MMPs as representative markers for ECM degradation, no anti-fibroblast directed therapy is currently available. Approaches to inhibit the joint destructive process in RA by elimination or inhibition of one proteolytic enzyme did not produce sufficient results in clinical trials regardless of supportive in-vivo results. (Schedel J, et al., Gene Ther 2004, 11(13):1040-1047; Lewis E J, et al., Br J Pharmacol 1997, 121(3):540-546; Brown P D, Expert Opin Investig Drugs 2000, 9(9):2167-2177).
The difference between the targeting strategies of the approaches described supra, and a FAP-α-specific targeting approach, results from the substantial potential of FAP-α as a specific marker for synovial fibroblasts in RA, as described in the Detailed Description which follows. Thus, an embodiment of the present invention includes a diagnostic methodology for determining presence of RA in a patient, involving assaying a synovial tissue sample taken from the patient for the expression of FAP-α, wherein the expression of FAP-α is indicative of presence of RA in the patient.
Another embodiment of the present invention includes a therapeutic methodology involving inhibition of FAP-α playing its role in tissue remodeling, as well as focusing antibody mediated cytotoxic activity on this synovial cell type to eradicate FAP-α's role in the joint destructive process. It has now been found that these deleterious cells can be identified via analysis of at least one marker on their cellular surfaces. Identification of this marker has therapeutic ramifications which are set forth in the disclosure which follows.
These, as well as other features of the invention, will be disclosed in greater detail in the Detailed Description which follows.